IgG/IgA Subclasses
Measurement of IgG Subclasses
IgG subclass serum concentrations in healthy individuals vary with age, sex, race, Gm allotype and exposure to acute and chronic infections. When developing assays to measure IgG subclasses it is necessary to consider all these factors to ensure that the assays will meet clinical requirements in terms of sensitivity, specificity, reproducibility and quality of performance.
Specificity
Monoclonal antibodies are known to be highly specific for the target antigen however they may exhibit an allotype bias. This could lead to non-detection of a deficiency or reporting of a false deficiency. High quality polyclonal antisera with no IgG subclass cross-reactivity is therefore more suitable for the reliable measurement of subclass concentrations. Binding Site assays utilise highly specific, affinity-purified, polyclonal antisera raised in sheep using patented technology.
Sensitivity
IgG subclass deficiencies, particularly in paediatric patients, may lead to antibody concentrations below the detection level of some standard assays. Sensitive assays are essential to allow reporting of actual values, enabling the laboratory to provide clinicians with accurate information. By coating specific antibodies onto latex particles, the immune complexes formed within the assay become large enough to be detected nephelometrically. Binding Site assays for IgG3 and IgG4 utilise latex-enhanced, liquid stable reagents for greater sensitivity.
Standardisation
Calibration of an assay against an internationally recognised reference preparation will ensure that the results remain accurate and consistent. The original reference material for IgG subclasses was the WHO immunoglobulin reference preparation WHO67/97. Values were first assigned to this material in 1971 but in 1985 a different set of values was assigned, leading to calibration differences between commercial IgG subclass reagent suppliers. In 1997 Carr-Smith et al assigned IgG subclass values to the international serum protein reference material CRM470. This material is available to purchase from The European Commission, Institute for Reference Materials and Measurements, Belgium and is the most commonly used reference material for commercial IgG assays. Binding Site IgG subclass assays are calibrated against CRM470. (Conversion factors are also supplied for customers wishing to use assays calibrated against the original WHO67/97.)
Quality
As the market leader in IgG subclass assays, Binding Site is acutely aware of the need for good quality control. Our laboratories are ISO9001:2000 and FDA certified, and we participate in several external quality assurance schemes. We also offer customers the opportunity to participate in our own IMMPROVE Subclass Quality Assurance Scheme.
Quality control procedures for all our IgG subclass products include extensive checking against internal reference materials and CRM470. Kits are CE marked for the majority of European countries, FDA cleared and have a minimum shelf life of 12 months from date of manufacture.
Measurement of IgA Subclasses
IgA is responsible for mucosal immunity and is therefore the first line of defence for the majority of infections. Deficiencies in IgA subclasses or alterations in the IgA1/IgA2 ratio have been related to specific diseases, recurrent infections and anaphylactic transfusion reactions. Normal serum concentrations of both IgA subclasses may be very low therefore a sensitive, reproducible assay is essential, particularly for paediatric samples. Binding Site IgA subclass assays also utilise highly specific high affinity, polyclonal antisera and latex - enhanced reagents enable accurate quantitation to very low levels.
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